Recombinant l-asparaginase production using Pichia pastoris (MUTs strain): establishment of conditions for growth and induction phases

Pillaca-Pullo O.; Rodrigues D.; Sánchez-Moguel I.; Lopes A.; Pimenta M.; Basi T.; Feitosa V.; Iris Zavaleta A.; Monteiro G.; Pessoa A., Jr.; Vitolo M. · Journal of Chemical Technology and Biotechnology (2021) · DOI: 10.1002/jctb.6540

Resumen

BACKGROUND: l-asparaginase (ASNase), a biopharmaceutical enzyme used in the treatment of childhood lymphoid malignancies, is commercially produced from Escherichia coli and Erwinia chrysanthemi. However, it causes severe adverse effects due to allergenic prokaryotic epitopes on the protein surface. ASNase II from Saccharomyces cerevisiae can be a promising alternative source of this enzyme. In this study, conditions to produce ASNase from S. cerevisiae expressed in Pichia pastoris have been investigated in shake flasks and 3 L-bioreactor. We evaluated if medium composition, concentration of carbon source (i.e. glycerol), growth time, concentration of inducer (i.e. methanol), temperature and initial pH influenced both biomass and ASNase expression. RESULTS: Biomass of around 53 g L–1 and ASNase volumetric activity of 710 U L–1 were achieved using the buffered glycerol-complex medium (BMGY) containing 40 g L–1 glycerol, with induction after 141 h using 3.0% (v/v) methanol, at 20 °C and initial pH 6.0. CONCLUSION: The experiments performed in shake flasks were scalable to a 3 L-bioreactor, suggesting that this bioprocess could be scaled-up for industrial production.

Instituciones: Universidad Nacional Mayor de San Marcos; Universidad Privada San Juan Bautista